Abstract: In-depth understanding of the characteristics of physical and chemical properties of materials, the abundance of pathogenic bacteria and the effect of disinfectant and lime on the causative organisms of cow mastitis during the production process of cow dung regeneration bedding can provide data support and theoretical basis for the rational application of cow dung regeneration bedding in farms. In this study, based on the process of converting cow manure into regenerated bedding in the drum aerobic fermentation process, the number of Escherichia coli, Streptococcus, Staphylococcus aureus and Klebsiella in different locations of the drums during the production process, as well as the physico-chemical characteristics of bedding materials, were measured and the optimal amount of disinfectant and lime added were determined by using the plate culture method. The results showed that there was no significant correlation between E. coli, Klebsiella and Streptococcus and the number of days, and the overall indicators of various pathogenic bacteria at the discharge port were lower than those at the inlet port, which indicate that the padding regeneration system of high-temperature fermentation in drum (60-75 ℃, more than 12 h) had a significant effect on the inhibition of pathogenic bacteria. After regeneration, the pH of the padding was in the range of 7.5-9.0, and the materials′ ash, total carbon, total nitrogen concentrations showed a decreasing trend at the discharge port, which was in line with the actual fermentation law; different disinfectants concentration treatment under the pathogenic bacteria inhibition law is consistent. Disinfectant powder and lime to a certain extent can be bacteria, Klebsiella and streptococcus reproduction to achieve the effect of inhibition of its growth, of which 3‰ (mass percentage) concentration of disinfectant inhibition of bacterial effect is the best. The results of the study can provide a scientific basis for the efficient resourcing of cattle manure recycled bedding and the safe use of farms.